Project Summary Cocaine addiction continues to be a significant public health problem for which there are currently no effective FDA-approved pharmacological treatments. Therefore, there is a clear need to identify novel neural mechanisms underlying cocaine craving and relapse in order to develop new pharmacotherapies to treat this disease. We have recently shown that central glucagon-like peptide-1 receptors (GLP-1Rs) play an important role in cocaine reinforcement and the reinstatement of cocaine seeking, an animal model of relapse. Specifically, we identified behaviorally relevant doses of a GLP-1R agonist that selectively reduced cocaine seeking and did not produce adverse effects commonly associated with these medications in humans and rodents. While these exciting findings clearly highlight a novel neuroendocrine mechanism that could be targeted to prevent cocaine craving-induced relapse, the neural mechanisms mediating the effects of GLP-1R agonists on cocaine seeking remain unclear. One goal of this proposal is to fill the gaps in our understanding of the central GLP-1 circuits regulating cocaine seeking. In Aim 1, we will extend this circuitry to include the lateral dorsal tegmental area (LDTg) and amygdala, two nuclei known to play critical roles in the reinstatement of cocaine seeking. We will also use a systems neuroscience approach to phenotype GLP-1R-expressing cells and identify their targets. Findings from these studies will provide the first comprehensive neuroanatomical map of GLP-1 circuits in the rat brain. Our pilot studies also reveal that GLP-1Rs are expressed on astrocytes and neurons in the rat brain. Using viral-mediated gene delivery and fiber photometry approaches in Aim 2, we will determine if reduced GLP-1R expression selectively on astrocytes and/or neurons prevents the suppressive effects of a GLP-1R agonist on cocaine seeking. In addition, we will investigate cell-type specific effects of GLP-1R activation on astrocyte activity and neuronal function during the reinstatement of cocaine seeking. We have also discovered that cocaine self-administration and subsequent abstinence dynamically regulate expression of endogenous preproglucagon (PPG), the gene that encodes GLP-1, in the hindbrain. These provocative findings suggest that reduced endogenous PPG expression during abstinence may facilitate cocaine seeking. However, the molecular and epigenetic mechanisms by which cocaine exposure regulates PPG expression are unknown. We will use chromatin immunoprecipitation (ChIP) methods in Aim 3 to identify the histone posttranslational modifications (PTMs) associated with reduced PPG transcription in the hindbrain. We will also identify transcription factors that regulate cocaine-induced changes in PPG mRNA expression. Together, these studies will provide new mechanistic insights into how cocaine exposure influences endogenous central GLP-1 signaling and highlight molecular substrates that could serve as targets for novel medications to treat cocaine addiction.